ABSTRACT In 2013 alone, ~230,000 children were infected with HIV worldwide, with most cases due to transmission to infants from their mother during pregnancy, birth, or breastfeeding. Early treatment can improve health outcomes, but most HIV-exposed infants do not get the testing needed to initiate therapy. The 2010 WHO guidelines recommended infant testing throughout the risk period from birth through breastfeeding. Existing tests typically require expensive instruments and trained staff. Disposable rapid tests (similar to pregnancy tests) are the only options available in the lowest-resource settings, but existing rapid HIV tests that work for diagnosis in adults fail completely for infant diagnosis due to the presence of maternal antibodies in the infant's blood. A rapid, low-cost, disposable infant HIV test would have immediate impact in the lowest-resources settings and provide a sustainable solution for the future. Specific requirements are a test that: 1) functions despite confounding maternal antibodies, and 2) addresses constraints (cost, ease-of-use, shelf stability) of the low-resource settings where infant diagnosis is most needed. Our goal is to develop an ultrasensitive signal- amplified test for diagnosis of infant HIV in a simple, rapid, disposable format (like a pregnancy test) appropriate for minimal-resource settings. Aim 1. Enable rapid test in low resource settings by developing a paper ELISA test that exhibits the same sensitivity as FDA-approved p24 ELISA. We will develop a simple HIV strip test with dry shelf-stable reagents that gives sensitivity comparable to existing lab tests. Aim 2. Enable a test for infant HIV diagnosis in low resource settings by developing an instrument-free module to isolate protein targets from whole blood. Detection of the protein biomarker requires removal of maternal antibodies prior to detection. We will develop a rapid paper module to capture and release the protein into a clean buffer for detection. Aim 3. Enable test-and-treat of HIV-infected infants in low-resource settings by demonstrating a rapid infant HIV test prototype that integrates sample processing and sensitive p24 detection in an easy-to-use disposable cartridge. User steps will be similar to existing rapid tests, and sample processing and ELISA will be done automatically.